Role of the C355 – C385 Disulfide Bonds in PACE4 Function

[摘要] Furin and PACE4 are part of a family of proteins called the proprotein convertases. Both proteins are critical for development, homeostasis, and disease pathology. Both proteins have two disulfide bonds within the catalytic domain. In furin, the bonds are C211-C260 and C303-C333, while the corresponding bonds in PACE4 are C263-C412 and C355-C385. PACE4 is like furin in structure and function, which will allow us to investigate any similarities that exist between the trafficking and function of these two proteins. Site-directed mutagenesis was performed to change the individual cysteine residues to serine residues. PACE4 constructs and a pro-vWF construct were used to perform a co-expression assay to examine if the disulfide bond knockout abolished the ability of PACE4 to process pro-vWF. RPE.40 cells were used, because they are furin null and do no produce PACE4 protein. Additionally, an activity assay was performed to further examine the effects of removing a disulfide bond from PACE4. Only the C355S PACE4 mutant was generated. Co-expression assays showed that the C355-C385 disulfide bond was critical for the ability of PACE4 to process substrates. This study allowed us to gain information about the disulfide bonds and PACE4 function, which will allow us to corroborate our findings about furin maturation.

[发布机构] University of Michigan

[效力级别] furin

[关键词] disulfide bond;furin;PACE4;proprotein convertase;protein maturation;protein trafficking;Molecular biology;Biology;Biochemistry;Biology